Hemoglobin is the oxygen-transport protein in the red blood cells. Hemoglobin in the blood carries oxygen from the respiratory organs (i.e. respiratory tract and lung) to organs and peripheral tissues to provide oxygen to the organs and the peripheral tissues and by doing so to assure normal physiological functions of the organs and the peripheral tissues.
In normal adult humans, hemoglobin is a hetero-tetramer, consisting of a pair of dissimilar subunits, including α1, α2, β1 and β2 subunits. While the backbone amino acid sequence determines the primary structure of each subunit, the intra-subunit hydrogen bonds and salt bridges formed within each of the subunits govern the secondary and tertiary structure of the subunits. Moreover, the inter-subunit hydrogen bonds and salt bridges formed between different subunits determine and regulate the quaternary structure of the tetrameric hemoglobin.
The quaternary structure of hemoglobin may exist in two allosteric conformation states, including a high oxygen affinity relaxed state (“R” state) and a low oxygen affinity tensed state (“T” state). Hemoglobin can bind oxygen and transform to the “R” state when transported to lungs where the partial pressure of oxygen PO2 is high, and release the bound oxygen to the organs and the peripheral tissues where the partial pressure of oxygen PO2 is low, and transform to the “T” state. A number of heterotropic effectors such as pH value, CO2 and 2,3-bisphosphoglycerate (2,3-BPG) play important roles in regulating the allosteric property of hemoglobin. Moreover, there are six inter-subunit hydrogen bonds in hemoglobin being capable of stabilizing hemoglobin in the low oxygen affinity “T” state, including α1Arg141---α2Asp126, α1Arg141---α2Lys127, α1Asp126---α2Arg141, α1Lys127---α2Arg141, β1His146---α2Lys40 and β2His146---α1Lys40. Among these six “T” state stabilizing inter-subunit contacts, four are related to αArg141 of hemoglobin, pointing to the crucial importance of this residue in sustaining the “T” state.
In general, hemoglobin with an impaired ability of carrying or releasing oxygen may cause a variety of syndromes such as anemia and dizziness; fatigue, weakness and shortness of breath are also frequently found in patients whose hemoglobin has defect oxygen-releasing capability. Syndromes, such as migraine, menstrual disorder and dysmenorrhea are also related with impaired oxygen-delivery efficiency of hemoglobin. Furthermore, insufficient oxygen uptake results in metabolism abnormality and dysfunction of the organs and the peripheral tissues, from which various diseases can begin to develop, including, but not limited to, hypertensions, cardiovascular and neurodegenerative diseases, and growth of carcinogenic cells. The conventional method broadly adopted to treat anemia involves transfusion of normal functional blood. However, this is a passive way of treatment and additional treatments must always be accompanied to alleviate the accompanying adverse side effects. For example, the iron-chelating agent must be applied to patients receiving the blood transfusion in order to down-regulate the iron level in blood to prevent iron-poisoning. In light of this, it is necessary to develop new strategies to improve the oxygen-releasing ability of hemoglobin to the organs and the peripheral tissues in human bodies and to treat various syndromes and diseases related with deficient oxygen delivery. Furthermore, many hemoglobin variants have reduced oxygen delivery capacity when compared with normal hemoglobin, which is often due to the altered allosteric properties or the loss of ability to interact with the endogenous allosteric effector, 2,3-bisphosphoglycerate as a result of structural modification. It is therefore also important to develop a new method to enhance the oxygen delivery ability of hemoglobin variants, recombinant hemoglobin or certain hemoglobin-based blood substitutes for the medical purposes.